Relationship between IgM antibody to human cytomegalovirus, virus load, donor and recipient serostatus, and administration of methylprednisolone as risk factors for cytomegalovirus disease after liver transplantation.

A retrospective study was performed on a selected cohort of 40 liver transplant recipients derived from the previous prospective follow-up of 162 liver transplant patients. The criterion for selection of this cohort was the presence of human cytomegalovirus (HCMV) DNAemia after transplantation, as determined by qualitative polymerase chain reaction (PCR). These 40 patients were followed longitudinally by quantitative PCR and by the new recombinant antigen-based AxSYM immunoassay for IgM to HCMV. The detection of IgM to CMV after transplantation was significantly associated with the development of HCMV disease in patients who had evidence of active HCMV replication in the blood by PCR (P=.01). On the basis of multivariate logistic regression analyses, the maximum titer of IgM detected after transplantation was a risk factor that was independent of augmented methylprednisolone and donor seropositivity. However, in multivariate analyses, elevated virus load continued to be the predominant risk factor for progression to HCMV disease.

Application of viral-load kinetics to identify patients who develop cytomegalovirus disease after transplantation.

BACKGROUND: Cytomegalovirus (CMV) continues to be a major problem post-transplantation; early markers for predicting patients at risk of CMV disease are needed. Peak CMV load in the blood correlates with CMV disease but frequently occurs too late to provide prognostic information. METHODS: 359 transplant recipients (162 liver, 87 renal, and 110 bone marrow) were prospectively monitored for CMV DNA in the blood with qualitative and quantitative PCR. 3873 samples were analysed. The CMV load in the first PCR-positive sample and the rate of increase in CMV load in blood during the initial phase of replication were assessed as risk factors for CMV disease using logistic regression. FINDINGS: 127 of the 359 patients had CMV DNA in the blood and 49 developed CMV disease. Initial viral load correlated significantly with peak CMV load (R2=0.47, p=<0.001) and with CMV disease (odds ratio 1.82 [95% CI 1.11-2.98; p=0.02; 1.34 [1.07-1.68], p=0.01, and 1.52 [1.13-2.05], p=0.006, per 0.25 log10 increase in viral load for liver, renal, and bone-marrow patients, respectively). The rate of increase in CMV load between the last PCR-negative and first PCR-positive sample was significantly faster in patients with CMV disease (0.33 log10 versus 0.19 log10 genomes/mL daily, p<0.001). In multivariate-regression analyses, both initial CMV load and rate of viral load increase were independent risk factors for CMV disease (1.28 [1.06-1.52], p=0.01, per 0.25 log10 increase in CMV load and 1.52 [1.06-2.17], p=0.02, per 0.1 log10 increase in CMV load/mL daily, respectively). INTERPRETATION: CMV load in the initial phase of active infection and the rate of increase in viral load both correlate with CMV disease in transplant recipients; in combination, they have the potential to identify patients at imminent risk of CMV disease.

Human herpesviruses 6 and 7 as potential pathogens after liver transplant: prospective comparison with the effect of cytomegalovirus.

Because cytomegalovirus (CMV) is an important opportunistic infection after liver transplant, we conducted a prospective study to see if the same applied to human herpesviruses (HHV)-6 and -7. We used polymerase chain reaction (PCR) methods optimised to detect active, not latent, infection and studied patients not receiving antiviral prophylaxis for CMV. Post-transplant, 536 blood samples were tested by PCR (median 7; range 4-50). Active infection with CMV was detected in 28/60 (47%), HHV-6 in 19/60 (32%), and HHV-7 in 29/60 (48%) of patients. The PCR-positive samples were tested by quantitative-competitive PCR to measure the virus load of each betaherpesvirus. The median peak virus load for CMV was significantly greater than that for HHV-6 or HHV-7. Detailed clinicopathological analyses for the whole population showed that CMV and HHV-6 were each significantly associated with biopsy-proven graft rejection. Individual case histories suggested that HHV-6 and HHV-7 may be the cause of some episodes of hepatitis and pyrexia. It is concluded that HHV-6 is a previously unrecognized contributor to the morbidity of liver transplantation, that HHV-7 may also be important and that both viruses should be included in the differential diagnosis of graft dysfunction.

The dynamics of human cytomegalovirus replication in vivo.

Cytomegalovirus (CMV) is generally described as a slowly replicating virus. During studies of immunocompromised patients, we observed rapid changes in the quantity of CMV DNA present in serial blood samples by quantitative-competitive polymerase chain reaction commensurate with a doubling time of <2 d. To further investigate the dynamics of replication in vivo, patients in three distinct situations were studied in detail: (a) those receiving intravenous ganciclovir; (b) those in whom ganciclovir-resistant strains appeared during long-term therapy; and (c) those in whom ganciclovir-resistant strains disappeared with alternative drug therapy. In all cases, it was possible to provide accurate estimates of the doubling time of CMV and its half-life of disappearance after antiviral chemotherapy. The results from all three approaches demonstrated that the doubling time/half-life of CMV in blood is approximately 1 d when frequent samples are collected. These results show that CMV DNA replication in vivo is a highly dynamic process. We conclude that the reputation of CMV as a slowly replicating virus based on the time taken to produce cytopathic effects in vitro is unwarranted. These findings have implications for the potency, dose, and duration of antiviral chemotherapy needed for the effective treatment of this important human pathogen.

Diagnostic approaches to cytomegalovirus infection in bone marrow and organ transplantation.

Cytomegalovirus (CMV) continues to be a clinical problem, impairing the overall success rate of transplantation, either through direct involvement of a variety of end-organs or by inducing indirect effects such as graft rejection. We review here how the virus manages to evade host immune responses and replicate extensively in allograft recipients. Recent studies show that the quantity of CMV (viral load) is related directly to the development of CMV disease. We review how clinically significant levels of CMV viral load can be defined and summarize the results of studies showing that a high CMV viral load is the major determinant of CMV disease, explaining the previously reported risk factors of pre-transplant serostatus and the post-transplant detection of CMV viremia.Note

Transcription of the human cytomegalovirus natural killer decoy gene, UL18, in vitro and in vivo.

Multiplex RT-PCR analysis of human cytomegalovirus (HCMV) replication in human fibroblasts showed transcription of the natural killer (NK) cell decoy gene, UL18, from 72 h onwards. Transcription of glycoprotein B (gpUL55; a late gene) occurred from early time-points and peaked at 24 h post-infection. UL18 mRNA was also detected in the peripheral blood mononuclear cells of organ transplant recipients with HCMV viraemia, especially those with HCMV DNA virus loads greater than 10(5) genomes/ml whole blood. Thus, UL18 is produced via a low abundance transcript late during the infectious cycle at a time coincidental with the increased risk of NK cell lysis as a consequence of class I HLA down-regulation.

Interrelationships among quantity of human cytomegalovirus (HCMV) DNA in blood, donor-recipient serostatus, and administration of methylprednisolone as risk factors for HCMV disease following liver transplantation.

Longitudinal analysis of 162 liver transplant recipients identified 51 patients who were viremic. Virus load was determined in 47 of these patients using quantitative-competitive polymerase chain reaction. Peak virus load was significantly higher in 20 symptomatic patients than 27 asymptomatic patients (P < .0001). Elevated virus load, donor seropositivity, and total methylprednisolone dosage were risk factors for human cytomegalovirus (HCMV) disease (odds ratio [OR], 2.22/0.25 log10 increase in virus load, P = .001; OR, 4.11, P = .05; OR, 1.30/1-g increment in methylprednisolone, P = .01). Methylprednisolone and virus load were independent risk factors in a multivariate analysis (OR, 2.70/1-g increase, P = .003; OR, 1.61/0.25 log10 increase, P = .03, respectively). Virus loads of 10(4.75)-10(5.25) genomes/mL of blood were associated with an increased disease probability; the latter was shifted to lower virus loads with increasing quantities of methylprednisolone. These data illustrate the central role of virus load in HCMV pathogenesis.

Quantity of cytomegalovirus viruria is a major risk factor for cytomegalovirus disease after renal transplantation.

Studies have shown that risk factors for human cytomegalovirus (HCMV) disease after renal transplant include primary infection (virus of donor origin infecting a non-immune individual), re-infection (virus of donor origin infecting a immune individual), and the detection of viraemia (as a marker of virus dissemination). We now report that viral load in the urine is also a significant factor in HCMV disease and is one of the main mechanisms underlying the risk associated with viraemia and donor serostatus. Longitudinal analysis of a group of 196 renal recipient identified 35 recipients who were PCR positive for HCMV in urine. Elevated viral loads were present in symptomatic patients, viraemic patients, and patients experiencing primary HCMV infection. Disease was associated with the peak quantity of virus present in the urine during the post-transplant period (P = 0.0001), with viraemia (P = 0.0003), and with transplantation of a seropositive donor (P = 0.03). Univariate logistic regression analysis showed that increases of 0.25 log10 in viral load were associated with a 179% increased risk of disease (odds ratio = 2.79; 95% C.I. 1.22-6.39; P = 0.02). This effect persisted in a multivariate logistic analysis when viraemia was incorporated (odds ratio = 2.77; 95% C.I. 1.07-7.18; P = 0.04). In contrast, the significant association between viraemia and disease observed in univariate analysis (odds ratio = 23.75; 95% C.I. 3.69-152.90; P = 0.0009) became marginally non-significant in multivariate analysis once viral load had been controlled for (odds ratio = 34.54; 95% C.I. 0.75-1599.00; P = 0.07). The computed probability of disease showed that a rapid transition occurred at viral loads between 10(5.7) and 10(6.5) genomes/ml urine in non-viraemic patients compared to viral loads between 10(5.0) and 10(5.7) genomes/ml urine in patients with concurrent viraemia. The implications of these findings for understanding HCMV pathogenesis, improving patient management, and optimising trials of antiviral treatment are discussed.